Fig. 1. Identification of CA125 as a counter receptor of galectin-1. (A) Affinity purification of galectin-1-interacting proteins. Both soluble (lanes 1, 2, 5, 6) and membrane (lanes 3, 4, 7, 8) fractions of HeLa cells were incubated with either GST—galectin-1 beads (lanes 2, 4, 6, 8) or GST beads as a control (lanes 1, 3, 5, 7). Bound proteins were eluted sequentially with lactose (lanes 1-4) and glutathione (lanes 5-8), followed by separation on Novex NuPage 10% Bis-Tris gels. Protein bands were visualized using SilverQuest. (B) Immunoblot analysis of the proteins eluted from the GST—galectin-1 and GST matrices, respectively. The fractions were loaded in the same order as shown in panel A. The anti-CA125 antibody OC125 was used as primary antibody followed by detection by electrochemiluminescence. (C) Immunoblot analysis as shown in panel B employing an anti-CA125-C-TERM_1-356 antiserum for the detection of CA125-derived fragments. (D) Amino acid sequence of CA125-C-TERM. Boxed sequences indicate tryptic peptides derived from band 1 (panel A, lane 2) as identified by mass spectrometry.

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